The Role of Ring Finger Protein 213 in Fatty Acid-induced Lipotoxicity

Abstract

Non-alcoholic steatohepatitis (NASH) is a progressive liver disease that manifests as liver inflammation and damage due to an excessive buildup of fat. NASH is increasing at an alarming rate worldwide and currently has an unmet medical need. Lipotoxicity is thought to be a leading cause of NASH, which can be recapitulated in cells by acute treatment with saturated fatty acids (SFA). However, mechanistic insights are lacking with respect to how cells adapt to lipotoxic stress. Here, I characterized the cellular and pathophysiologic function of Ringer Finger 213 (RNF213), which our lab identified in a genome-wide short-hairpin RNA-based screen for genetic factors controlling cellular response to lipotoxicity. The central hypothesis is that the deficiency of RNF213 promotes fatty acid desaturation to neutralize lipotoxic stress in cells and in vivo. I took the gene knockout approach to test this hypothesis in cells and a murine NASH model. RNF213 knockout (KO) prevents palmitate-induced cell death despite gaining an increased capacity of unfolded protein response in MEFs. Global lipidomic profiling revealed that RNF213 KO restores lipotoxic species such as disaturated DAGs in cellular lipidome to control levels, normalizes fatty acid desaturation and reduces DAG biosynthesis upon palmitate treatment in MEFs. This is associated with increased enzyme activity of stearoyl-CoA desaturase, but how RNF213 regulates SCD1 activity remains unknown. To explore the cell biology of RNF213, I collaborated with Dr. Ayon Ibrahim to utilize a fluorescent-tagged knock-in cell line and found that endogenous RNF213 translocates partially to the ER membrane upon palmitate treatment but not oleic acid in SUM159 cells. Next, I examined the liver phenotype of RNF213 KO mice in a dietary NASH model. RNF213 KO mouse livers show a mild protection phenotype against liver damage and does not alter hepatic steatosis. Global lipid profiling reveals that RNF213 KO does not affect fatty acid desaturation in mouse livers. These findings demonstrate that RNF213 is a promoter of cellular lipotoxicity with an unknown mechanism, which is associated with fatty desaturation. However, the data did not favorably support the role of RNF213 in NASH in the context of saturated fatty acid toxicity in vivo, which may be due to the diet not raising levels conducive for severe NASH development. For future cellular screening, utilization of primary hepatocytes or liver organoids would be crucial for elucidating the translational potential of RNF213 and its potential in therapy for metabolic diseases.